Cellular interactions between hematopoietic stem and progenitor cells (SC/Ps) and the hematopoietic microenvironment (HM) play a critical role in regulating hematopoiesis. At least three classes of molecules contribute to the maintenance of hematopoiesis. Cytokines and chemokines stimulate the survival, growth, and migration of SC/Ps. Adhesion molecules (AMs) support the physical association of SC/Ps within the hematopoietic niches and regulate growth, homing and mobilization of SC/Ps. Nevertheless, little is known about the signaling mechanisms that control SC/P functions downstream from these molecules. Identification of key intracellular signaling molecules and pathways involved in regulating SC/P functions will facilitate the design of specific molecular therapies for the treatment of malignancies involving defects in SC/P function. Our long-range goal is to understand signaling mechanisms that control growth and actin-based functions in SC/Ps. Understanding these mechanisms is critical to the biology of SC/P amplification, homing and mobilization for translational applications of transplantation and gene therapy of blood disorders. The objective of this application is to determine how Src family of signaling molecules regulate growth and actin-based functions in c-Kit+Scal+ Lin- cells. The central hypothesis of the application is that hematopoietic specific Src family kinase (SFK) possess both unique as well as overlapping mechanisms to regulate growth and actin-based functions in c- Kit+Sca-l+Lin- cells. Among the known SFKs, three highly homologous SFKs Lyn, Hck, and Fgr are expressed only in SC/Ps. Amino acid sequence differences in these molecules are restricted predominantly to the amino terminus region. Despite their sequence similarities, we will present preliminary data to support our hypothesis that SFKs differentially (positively and negatively) regulate the magnitude and duration of cytokine-mediated growth in SC/Ps in part via regulation of phosphatase activation. The SC/P phenotypic changes we have identified associated with Lyn-deficiency are apparent in spite of normal Hck and Fgr expression. How these hematopoietic SFKs differentially (positively and negatively) modulate SC/P growth and survival given their limited amino acid differences will constitute a focus of this proposal. Our preliminary observations also lead us to hypothesize that SFKs regulate actin-based functions in SC/Ps, including defective chemotaxis in vitro and redistribution in vivo of c-Kit+Lin-Sca-l+ cells. Our proposed studies will provide unique insights into the physiologic significance of Lyn, Hck, and Fgr SFKs in the regulation of SC/P growth and actin-based functions.